SYBR Safe™ versus
ethidium bromide
Nikita Warner, Ph.D.
Assoc. Product Manager
Nucleic Acid Separation & Analysis
What is SYBR Safe™?
A DNA gel stain that is:
• Better for you
– displays no acute oral toxicity and greatly reduced genotocixity
• Better for the environment
– offers non-hazardous waste status and Clean Water Act
compliance
• Better for your institution
– reduces waste storage and disposal costs
• Better for your sample
– improves cloning efficiency when used with blue-light illumination
without compromising detection sensitivity as compared to
ethidium bromide
SYBR Safe™ is a better DNA gel stain
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How is SYBR Safe™ better for me?
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Oral toxicity evaluation
Analysis
Method
Results
Aquatic toxicity*
Fathead minnow CA Title Not hazardous or toxic to
22 acute screening
aquatic life
Oral toxicity**
EPA Acute Oral Toxicity
Test OPPTS 870.1100
LD50 > 5000 mg/kg
* Performed by AMEC Earth and Environmental San Diego Bioassay
Laboratory, San Diego, CA.
** Peformed by Northview Pacific Laboratories, Inc., Hercules, CA.
Displays no acute oral toxicity
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Genotoxicity studies
In Vitro
Test*
Cell Type
Result with S9
Activation
Result without S9
Activation
Transformation
Syrian hamster embryo
(SHE) cells
NA
Negative
Chromosomal
aberrations2
Cultured human peripheral
blood lymphocytes
Negative
Negative
Forward
mutation3,4
L5178YTK+/- mouse
lymphoma cells
Negative
Negative
1
* In vitro tests were performed by Covance Laboratories Inc., Vienna, VA.
Mammalian
S9 fraction obtained from Aroclor 1254 induced rat liver. NA = Not applicable.
1Fundamental and Molecular Mechanisms of Mutagenesis 356, 1 (1996); 2Evans, H.J., in
Chemical Mutagens, Principles and Methods for Their Detection, A. Hollaender, Ed., Vol.
4, (1976) pp. 129; 3Mutation Res 72, 447 (1980); 4Mutation Res 59, 61 (1979).
Displays greatly reduced genotocixity
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Ames test results
Causes fewer mutations in the standard Ames test
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How is SYBR Safe™ better for the environment?
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Hazardous waste testing
According to the guidelines of the EPA, SYBR Safe™ DNA gel stain is not
• corrosive
• ignitable, or
• reactive
Analysis*
Method
Results
Corrosivity
EPA 150.1
Not corrosive (pH = 8.25)
Corrosivity (by
Corrositex)
DOT-E 10904
Category 2 noncorrosive
Ignitability
EPA 1010
Not ignitable (<212°F)
Reactivity
EPA 9010B/9030A
No reactivity detected
* All tests were performed by AMEC Earth and Environmental San Diego
Bioassay Laboratory, San Diego, CA.
Not classified as hazardous waste under U.S. federal regulations
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NPDES analysis
Tests performed according to National Pollutant Discharge Elimination System
(NPDES) guidelines indicate that SYBR Safe™ DNA gel stain does not differ
significantly from 0.5X TBE buffer
Test
SYBR Safe Stain in 0.5X TBE
0.5X TBE
PH (150.1)
8.45
8.48
Total cynaide (335.2)
None detected
None detected
Chemical oxygen demand (COD; 410.1)
7020
6840
Ammonia as nitrogen (350.1)
253
248
Total organic carbon (415.1)
2480
2360
Total phenoloics (420.1)
None detected
None detected
Organochlorine pesticides and PCBs (608M)
None detected
None detected
Semi-volatile organic compounds (625)
None detected
None detected
Volatile organic compounds (624)
None detected
None detected
Priority Pollutant Metals (Sb, As, Be, Cd, Cr,
Cu, Pb, Hg, Ni, Se, Ag, Tl, Zn) (EPA
200.7/200 series)
None detected
None detected
* All tests were performed by Columbia Analytical Services, Inc., Kelso, WA. Methods used were as outlined in the Code of Federal
Regulations (CFR) Title 40, Part 136.
SYBR Safe™ DNA gel stain does not differ significantly from 0.5X TBE buffer 9
Organic compound analysis
Tested were performed on 0.5X TBE and 1X SYBR Safe™ in 0.5X TBE for:
• Organochlorine pesticides and PCBs (608M)
• Volatile organic compounds (624)
• Semi-volatile organic compounds (625)
None of the organic pollutant compounds tested for were detected in
either sample
SYBR Safe™ DNA gel stain in 0.5X TBE is indistinguishable from 0.5X TBE alone
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How is SYBR Safe™ better for my institution?
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SYBR Safe™ Institutional Savings
Case Study: Forsyth Institute, Boston, MA (data provided by Forsyth Institute)
Ethidium bromide
SYBR Safe™
Reagent Costs
•10 ml of 10 mg/ml concentrate @ $32 per bottle
•5 µl concentrate per gel
•$0.016 per gel
Disposal costs - Materials
•20 L capacity charcoal filter @ $33 per filter
•0.1 L per gel
•$0.165 per gel
Disposal costs - Time & Labor
•4 hours labor @ $20 per hour to filter 20 L of
waste
•0.1 L of waste per gel
•$0.40 per gel
Overhead disposal costs ++
Water usage (suction for filtration)
Hazardous waste fees (for filter disposal)
Additional uncalculated costs
Total ethidium bromide cost
$0.58++ per gel
Reagent Costs
•400 µl concentrate @ $40 per bottle
•5 µl concentrate per gel
•$0.50 per gel
Disposal costs – Materials
•$0.00
Disposal costs - Time & Labor
•$0.00
Overhead disposal costs
•None
•Total SYBR Safe™ cost
•$0.50 per gel
For more details, visit:
www.invitrogen.com/sybrsafe
14% institutional savings at a Boston institute
Invitrogen Proprietary and Confidential
SYBR Safe™ Waste Disposal at ISU
Iowa State University, Ames IA
• Wastewater officials in Ames, Iowa approved the disposal of SYBR Safe™ as nontoxic waste
• Diluted dye solution can be poured directly down the drain at Iowa State University
ISU scientists can dispose SYBR Safe™ liquid waste down the drain
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How is SYBR Safe™ better for my sample?
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Sensitivity comparison
Kb
23.1
9.4
6.6
4.4
2.3
2.0
Ethidium bromide
SYBR Safe™
Marker is l DNA/Hind III Fragment
SYBR Safe™ has comparable sensitivity to ethidium bromide
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E-Gel® - EtBr versus SYBR Safe™
1.2% E-Gel® with
ethidium bromide
0.3 second exposure
Same samples and volumes loaded on both gels. Both gels viewed on UV transilluminator
1.2% E-Gel® with
SYBR Safe™
1 second
exposure
Same sensitivity as ethidium bromide, but longer exposure
Invitrogen Proprietary and Confidential
How does one use SYBR Safe™?
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Using SYBR Safe™
Formats offered:
• Offered as 10,000X concentrate, or
• Ready-to-use solution in 1X TAE
• Ready-to-use solution in 0.5X TBE
• E-Gel® pre-cast agarose gels incorporated with SYBR Safe™
How to use:
• Incorporate into the gel, or
• Perform post-run staining, or
• Use E-Gel® with SYBR Safe™
SYBR Safe™ is convenient to use
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What does UV exposure do to DNA?
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Effect of UV on DNA
SYBR SafeTM & Blue Light
Time (Min)
2
4
8
16
Ethidium bromide & UV
Light
2
4
8
16
Nicked DNA
Supercoil DNA
CFUs obtained from
transformation with
constructs containing
inserts stained with
SYBR Safe™ or ethidium
bromide and viewed on
blue light or UV respectively
UV damages DNA and diminishes cloning results
Invitrogen Proprietary and Confidential
Eliminate the dangers of UV transillumination
Safe Imager Blue light Transilluminator
• Does not damage your skin and eyes
• Produces bright light and uniform emission
• Provides optimal excitation for SYBR Safe™ DNA gel stain
• For more information, visit www.invitrogen.com/safeimager
Optimum excitation of SYBR Safe™ and maximum safety
Invitrogen Proprietary and Confidential
Safe Imager versus the competition
Competitor blue light box
Safe Imager™ technology
Even illumination with the Safe Imager™
Invitrogen Proprietary and Confidential
Safe Imager for visualizing stained proteins
SYPRO Ruby staining
ng protein per band (approx.)
2x dilution series Mark 12 Markers
SafeImager
1 ng sensitivity with SYPRO Ruby and SafeImager
Invitrogen Proprietary and Confidential
Mark12 standards
0.125
0.25
0.5
1
2
4
8
16
32
64
128
256
500
Safe Imager and SYPRO-Ruby Staining of 2-D gel
Jurkat cell lysate
Avoid UV exposure when performing 2-D electrophoresis
Invitrogen Proprietary and Confidential
Lucy 565 (UV)
0.125
0.25
0.5
1
2
4
8
16
32
64
128
256
0.125
0.25
0.5
1
2
4
8
16
32
64
128
256
Safe Imager and SYPRO Orange protein stain
SYPRO Orange (Safe Imager)
Superior sensitivity with SYPRO Orange and Safe Imager versus Sigma’s Lucy 565
Invitrogen Proprietary and Confidential
Lucy 506 (Safe Imager)
0.125
0.25
0.5
1
2
4
8
16
32
64
128
256
0.25
0.125
0.5
1
2
4
8
16
32
64
128
256
Safe Imager and SYPRO Ruby protein stain
SYPRO Ruby (Safe Imager)
Mark12 standards
Superior sensitivity with SYPRO Ruby and Safe Imager versus Sigma’s Lucy 506
Invitrogen Proprietary and Confidential
The complete safety solution
SYBR Safe™ DNA Gel Stain
• Better for you
• Better for the environment
• Better for your institution
• Better for your sample
All this with …
• No compromise on performance
• For more information, visit
www.invitrogen.com/sybrsafe
Safe Imager Blue light Transilluminator
• Avoid UV exposure
• Get improved cloning efficiency
• Can be used with DNA and protein stains
SYBR Safe™ and Safe Imager™ offer the compete safety solution
Invitrogen Proprietary and Confidential
Information
Learn more about:
SYBR Safe™ at www.invitrogen.com/sybrsafe
Safe Imager™ at www.invitrogen.com/safeimager
Additional information or questions:
E-mail sybrsafe@invitrogen.com
We like you just the way you are!
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